For normal biological mineral deposition to proceed, a tight balance is required between the levels of extracellular inorganic phosphate (Pi and pyrophosphate (PPi)) The periodontium, the collection of upportive structures of the tooth root, is especially sensitive to Pi/PPi balance, as evidenced by dramatic cementum phenotypes resulting from mutations that alter local levels of Pi and PPi. A markedly increased cementum was demonstrated on the root surfaces of fully developed teeth in mice presenting mutations in either the multiple-pass transmembrane ankylosis protein (ANK) or plasma cell membrane glycoprotein-1 (PC-1); both mutations result in reduced extracellular PPi though via different mechanisms. Conversely, mice and humans with defective tissue nonspecific alkaline phosphatase (TNAP) present locally increased levels of PPi, leading to defective formation of acellular cementum, and subsequently, premature tooth loss. In contrast to these cementum phenotypes, the underlying mineralized tissue, dentin, seems to develop normally in conditions of altered Pi/PPi. Based on these observations, we hypothesize that periodontal ligament (PDL) cells adjacent to the tooth root cementum are more sensitive to changes in Pi levels than pulp cells adjacent to the dentin. In order to resolve the differential regulation of these two mineralized tissues, the following aims are set: (1) To determine basal differences in gene/protein expression in pulp vs. PDL cells in healthy and hypophosphatasia-diagnosed (HPP) subjects, focusing on genes/proteins associated with differentiation/mineralization and Pi/PPi metabolism; (2) To determine the effect that Pi treatment exerts on pulp vs. PDL cells, in vitro; and (3) To demonstrate that knocking down TNAP function in pulp and PDL cells harvested from healthy subjects results in a more dramatic change in PDL vs. pulp cell behavior, in vitro. Establishing the mechanisms by which Pi/PPi influence the periodontium is a critical step towards understanding the development of these tissues, with the ultimate goal of applying further insight to the design of therapies to regenerate periodontal tissues. [unreadable] [unreadable] [unreadable]